PROJECT SUMMARY While significant evidence has demonstrated that obesity increases the risk of metastasis, the molecular mechanisms by which obesity contributes to the metastatic progression of breast cancer are unclear. Further, recent research in cancer development and progression has highlighted the role of metabolic reprogramming, which results in an increased supply of the cellular building blocks necessary for the increased cell proliferation and in adaptations required for cell survival in changing nutrient- and oxygen-containing environments. Research from our team and others demonstrates that the metabolic enzyme, pyruvate carboxylase, is upregulated during obesity and that this upregulation correlates strongly with breast cancer progression. Additional studies suggest that leptin, an adipokine whose expression is increased during obesity and whose receptor's expression is enhanced in metastatic cells, drives pyruvate carboxylase expression in breast cancer cells. Importantly, recent studies demonstrate that genetic depletion of pyruvate carboxylase drastically inhibits breast cancer metastasis in several syngeneic mouse models. Despite the supporting evidence that pyruvate carboxylase contributes to breast cancer metastasis under obese conditions, the mechanisms by which this enzyme exerts this effect remain poorly understood. In the proposed studies, the research team will evaluate the mechanistic basis by which pyruvate carboxylase regulates obesity-driven breast cancer metastasis. They will test the hypotheses that leptin increases pyruvate carboxylase expression in mammary tissue during obese states, and that pyruvate carboxylase is critical for both migration and survival of extracellular matrix detachment, providing metabolic flexibility (e.g., glucose utilization and fatty acid metabolism) during metastasis. These hypotheses will be tested through completion of the following aims: 1) define the mechanisms of PC expression during metastasis; 2) elucidate the metabolic mechanisms by which PC promotes metastasis; and 3) establish the mechanisms by which leptin-regulated PC expression contributes to obesity-driven metastasis. Completion of these studies will result in valuable mechanistic information that could guide future development of evidence-based recommendations for those who are overweight and obese and that will help reduce breast cancer metastasis.